Protection of crops against fungal pathogens is one of the most significant unmet needs in agriculture. Despite these significant losses, less than 5 percent of U.S. corn and soybean acreage is treated with fungicides (Gianessi and Marcelli (2000) Pesticide Use in U.S. Crop Production: 1997, National Summary Report, November, 2000), for agronomic reasons and due to the diverse nature of the pathogens responsible for those losses.
In conventional pathogen-resistant crop varieties, resistance is achieved by using standard breeding techniques to introgress resistance (R) genes, which recognize or interact with pathogen virulence factors and activate defense responses, from wild germplasm into domesticated germplasm. However, R gene-mediated resistance is not usually durable because the pathogen mutates, eliminating the virulence factor detected by the plant. Since virulence factors appear to have redundant functions, individual factors can be lost with little, if any, diminished pathogenicity. Only in a few rare cases is durable resistance observed, and this is usually attributed to an essential function of a given virulence factor in the host-pathogen interaction. Moreover, and very importantly, R gene-medicated resistance protects crops against a limited spectrum of fungal pathogens. Most crops suffer from multiple pathogen problems, so that the industry seeks broad-spectrum, durable disease solutions.
The expression of the defense response can be engineered by altering the expression of regulatory proteins such as transcription factors (reviewed in Gurr and Rushton (2005) Trends Biotechnol. 23: 275-282). We have previously shown that constitutive and ectopic overexpression of key transcription factors involved in the natural defense response results in enhanced disease resistance in transgenic plants (e.g., see U.S. Pat. No. 6,664,446 or US Patent Application 20030046723). In many instances, the gain of function phenotype (disease resistance) is observed in interactions with multiple fungal and bacterial pathogens, a major advantage for the engineering of this trait in crops. This provides experimental evidence that altering the expression of natural defense responses is an effective method for engineering disease resistance in plants.
The applicability of this technology to crop species may be limited by negative side effects associated with constitutive overexpression of disease defense protein(s). Pleiotropic effects such as delayed growth and development and alteration in flowering time are common. It has been proposed that genes conferring resistance to pathogens impose a cost on overall fitness and development. Plants have achieved a balance between fitness and resistance by the evolution of inducible defenses.
The development of effective resistance of crops to different classes of pathogens will require the dissociation of the gain of function phenotype (disease resistance) from the negative side effects. We hypothesize that limiting overexpression of disease resistance transcription factors to infected tissues, only when disease pressure arises, will significantly reduce or eliminate the impact on yield and fitness, while retaining the gain of function phenotype. The present invention addresses the difficulties in identifying promoters with unique expression characteristics for applicability in the development of disease resistance in crops. We believe that the solution to this technical problems lies with the selection of plant promoters with key expression characteristics. These promoters may also be useful for controlled expression of other defense regulatory proteins, antimicrobial proteins, elicitors that induce defense responses, etc.